Hannes Koppel

Anserine inhibits carnosine degradation but in human serum carnosinase (CN1) is not correlated with histidine dipeptide concentration

BACKGROUND We reported an association of a particular allele of the carnosinase (CNDP1 Mannheim) gene with reduced serum carnosinase (CN1) activity and absence of nephropathy in diabetic patients. Carnosine protects against the adverse effects of high glucose levels but serum carnosine concentration was generally low. METHODS We measured the concentration of two further histidine dipeptides, anserine and homocarnosine, via HPLC. CN1 activity was measured fluorometically and for concentration we developed a capture ELISA. RESULTS We found an association between the CNDP1 Mannheim allele and reduced serum CN1 activity for all three dipeptides but no correlation to serum concentrations although anserine and homocarnosine inhibited carnosinase activity. Patients with liver cirrhosis have low CN1 activity (0.24 ± 0.17 μmol/ml/h, n=7 males; normal range: 3.2 ± 1.1, n=104; p<0.05) and CN1 concentrations (2.3 ± 1.5 μg/ml; normal range: 24.9 ± 8.9, p<0.05) but surprisingly, histidine dipeptide concentrations in serum are not increased compared to controls. CONCLUSIONS Serum histidine dipeptide concentrations are not correlated to CN1 activity. The protective effect of low CN1 activity might be related either to turnover of CN1 substrates or a protective function of dipeptides might be localized in other tissues.

L-carnosine inhibits high-glucose-mediated matrix accumulation in human mesangial cells by interfering with TGF-β production and signalling

BACKGROUND Transforming growth factor beta is recognized as a major cytokine in extracellular matrix (ECM) pathobiology as occurs in diabetic nephropathy. While experimental studies have advanced a protective role of carnosine for diabetic complications, a link between carnosine, TGF-β and matrix accumulation remains to be elucidated. In the present study, we tested the hypothesis that L-carnosine inhibits TGF-β production and signalling, thereby reducing hyperglycaemia-associated ECM accumulation. METHODS Human mesangial cells (MC) were cultured in high-glucose (HG, 25 mM D-glucose) medium alone or in HG medium to which 20 mM L-carnosine was added. Collagen VI (Col6) and fibronectin (FN) deposition and messenger RNA expression were studied. In addition, TGF-β production and activation of Smad1/5/8 (ALK1) and Smad2/3 (ALK5) pathways were assessed. RESULTS Under HG conditions, deposition of Col6 and FN were increased 1.4- and 1.6-fold. This was significantly inhibited on the protein and messenger RNA level by L-carnosine. TGF-β production increased under HG conditions but was completely normalized by addition of L-carnosine. Addition of exogenous TGF-β could not overcome the effect of L-carnosine on Col6 and FN expression, indicating additionally interference with TGF-β downstream signalling. Along the same line, L-carnosine reduced TGF-β-mediated Smad2 phosphorylation, suggesting an inhibitory effect on ALK5 signalling. ALK1 signalling remained unchanged. Under HG conditions, pharmacologic inhibition of ALK5 prevented Col6 accumulation but did not change FN deposition. CONCLUSIONS L-carnosine can modulate matrix accumulation in two ways. Firstly, inhibition of TGF-β production might result in an overall inhibition of matrix accumulation and secondly, L-carnosine inhibits TGF-β-induced matrix accumulation, most likely via inhibition of the ALK5 pathway.

Carnosine treatment in combination with ACE inhibition in diabetic rats

In humans, we reported an association of a certain allele of carnosinase gene with reduced carnosinase activity and absence of nephropathy in diabetic patients. CN1 degrades histidine dipeptides such as carnosine and anserine. Further, we and others showed that treatment with carnosine improves renal function and wound healing in diabetic mice and rats. We now investigated the effects of carnosine treatment alone and in combination with ACE inhibition, a clinically established nephroprotective drug in diabetic nephropathy. Male Sprague-Dawley rats were injected i.v. with streptozotocin (STZ) to induce diabetes. After 4 weeks, rats were unilaterally nephrectomized and randomized for 24 weeks of treatment with carnosine, lisinopril or both. Renal CN1 protein concentrations were increased under diabetic conditions which correlated with decreased anserine levels. Carnosine treatment normalized CN1 abundance and reduced glucosuria, blood concentrations of glycosylated hemoglobin (HbA1c), carboxyl-methyl lysine (CML), N-acetylglucosamine (GlcNac; all p<0.05 vs. non-treated STZ rats), reduced cataract formation (p<0.05) and urinary albumin excretion (p<0.05), preserved podocyte number (p<0.05) and normalized the increased renal tissue CN1 protein concentration. Treatment with lisinopril had no effect on HbA1C, glucosuria, cataract formation and CN1 concentration, but reduced albumin excretion rate more effectively than carnosine treatment (p<0.05). Treatment with both carnosine and lisinopril combined the effects of single treatment, albeit without additive effect on podocyte number or albuminuria. Increased CN1 amount resulted in decreased anserine levels in the kidney. Both carnosine and lisinopril exert distinct beneficial effects in a standard model of diabetic nephropathy. Both drugs administered together combine the respective effects of single treatment, albeit without exerting additive nephroprotection.

In Others' Shoes: Do Individual Differences in Empathy and Theory of Mind Shape Social Preferences?

Abundant evidence across the behavioral and social sciences suggests that there are substantial individual differences in pro-social behavior. However, little is known about the psychological mechanisms that underlie social preferences. This paper investigates whether empathy and Theory of Mind shape individual differences in pro-social behavior as conventionally observed in neutrally framed social science experiments. Our results show that individual differences in the capacity for empathy do not shape social preferences. The results qualify the role of Theory of Mind in strategic interaction. We do not only show that fair individuals exhibit more accurate beliefs about the behavior of others but that Theory of Mind can be effectively used to pursue both self-interest and pro-social goals depending on the principle objectives of a person.

The CNDP1 (CTG)5 Polymorphism Is Associated with Biopsy-Proven Diabetic Nephropathy, Time on Hemodialysis, and Diabetes Duration

Considering that the homozygous CNDP1 (CTG)5 genotype affords protection against diabetic nephropathy (DN) in female patients with type 2 diabetes, this study assessed if this association remains gender-specific when applying clinical inclusion criteria (CIC-DN) or biopsy proof (BP-DN). Additionally, it assessed if the prevalence of the protective genotype changes with diabetes duration and time on hemodialysis and if this occurs in association with serum carnosinase (CN-1) activity. Whereas the distribution of the (CTG)5 homozygous genotype in the no-DN and CIC-DN patients was comparable, a lower frequency was found in the BP-DN patients, particularly in females. We observed a significant trend towards high frequencies of the (CTG)5 homozygous genotype with increased time on dialysis. This was also observed for diabetes duration but only reached significance when both (CTG)5 homo- and heterozygous patients were included. CN-1 activity negatively correlated with time on hemodialysis and was lower in (CTG)5 homozygous patients. The latter remained significant in female subjects after gender stratification. We confirm the association between the CNDP1 genotype and DN to be likely gender-specific. Although our data also suggest that (CTG)5 homozygous patients may have a survival advantage on dialysis and in diabetes, this hypothesis needs to be confirmed in a prospective cohort study.

Mixed messages on systemic therapies for diabetic retinopathy

The ACCORD study (Aug 7, p 419) has raised more questions about intensive systemic medical therapy in the management of diabetic retinopathy. First, why did intensive glycaemic control not prevent the primary composite microvascular outcome in the overall study, but slow progression of diabetic retinopathy in the ACCORD-Eye substudy? Second, how do clinicians balance the benefi ts of intensive glycaemic control in ACCORD with increased mortality? Third, ACCORD did not fi nd intensive blood pressure control to be useful in reducing progression of diabetic retinopathy. This fi nding contrasts with that of the UKPDS, although the targets for both intensive and standard control (systolic blood pressure <120 mm Hg and <140 mm Hg, respectively) in ACCORD were lower than in the UKPDS intensive control group (systolic blood pressure <150 mm Hg). Does this fi nding suggest a “fl oor” to benefi ts of lowering blood pressure below normal ranges? Finally, the implications of the impressive eff ects of fenofi brate on progression of diabetic retinopathy remain unclear. Although the FIELD study found that fenofi brate reduced the need for laser treatment, the results were limited by frequent, uneven statin use. ACCORD-Eye, which compared fenofi brate with placebo in patients already using statins, confi rmed the superiority of feno fi brate in preventing progression of diabetic retinopathy, with a risk reduction similar to that of FIELD (30–40%). However, as in FIELD, this eff ect seemed independent of lipid con centrations. Do fenofi brates act via novel therapeutic mechanisms, perhaps involving infl ammatory or endothelial dysfunction pathways? These fi ndings suggest that much remains to be learnt about the pathogenesis of diabetic retinopathy and the eff ectiveness of systemic therapy. How fi ndings from ACCORD, and other recent trials, can be applied to routine clinical practice needs clarifi cation.

L-Carnosin verhindert die Akkumulation von extrazellulärer Matrix unter HochGlucose über Modifikation des TGF-β Systems

Fragestellung: Die Vulnerabilitat von Diabetikern eine diabetische Nephropathie (DN) zu entwickeln ist u.a. genetisch festgelegt. Wir konnten das Gen CNDP-1 (Serum Carnosinase) identifizieren, dessen Polymorphismus im Signalpeptid mit der Empfanglichkeit fur DN assoziiert ist. Patienten, die homozygot fur die kurzeste allelische Form (Mannheim Allel) des Gens sind, entwickeln seltener eine diabetische Nephropathie. Diese allelische Form geht mit niedriger Aktivitat des Enzyms einher und fuhrt somit zu hoheren Carnosin (L-Carn) Spiegeln. Durch welchen Mechanismus L-Carn protektiv wirkt ist bisher nur unzureichend geklart. Das Signalmolekul TGF-β spielt bei der Entwicklung der DN eine Schlusselrolle. Es verursacht u.a die Akkumulation von extrazellularer Matrix (EZM), einem der Merkmale der DN. In der gegenwartigen Studie untersuchen wir, ob L-Carn in humanen Mesangialzellen (MC) Einfluss auf TGF-β und seine Signalwege hat. Methodik: Humane MC wurden fur 14 Tage in Standardmedium±25mM Glucose±20mM L-Carnosin gezuchtet. In den Zelluberstanden wurden die TGF-β Konzentrationen mittels Bioassay gemessen. Des Weiteren wurde die Produktion von EZM (Fibronektin (FN) und Collagen VI (Col6)) durch quantitative PCR (LightCycler) bestimmt. Die Akkumulation der EZM wurde uber indirekte Immunfluoreszenz bestimmt. Die TGF-β Signalwege wurden mithilfe von Western Blot des ALK1- (pSmad 1/5/8) und des ALK5 (pSmad 2) Signalweges untersucht. Ergebnisse: HochGlucose induziert die FN- und Col6-Ablagerung um den Faktor 1,4 bzw. 1,6. Dieser Effekt konnte durch den Zusatz von L-Carn vollstandig verhindert werden. Die Beobachtungen konnten auf mRNA Ebene bestatigt werden. Unter HochGlucose stieg die TGF-β Konzentration der Uberstande von 1,5 auf 2,0ng/ml/10E6 Zellen an. Zusatz von L-Carn normalisierte die Werte (1,5ng/ml/10E6 Zellen; p<0,001). Ebenso konnte die Akkumulation von FN und Col6 unter direkter Stimulation mit TGF-β durch die Zugabe von L-Carn gehemmt werden. Die Stimulation unter HochGlucose gezuchteter MC mit TGF-β zeigte eine Aktivierung sowohl von pSmad 1/5/8 als auch von pSmad 2, wobei die pSmad 2 Aktivierung uberwiegte. L-Carn reduzierte hierbei die Aktivierung von pSmad 2. Die exogene Inhibierung des ALK5 Signalwegs verhinderte die Akkumulation von Col6. Schlussfolgerungen: Unsere Daten zeigen, dass L-Carn die Akkumulation der extrazellularen Matrix auf zwei Wegen verhindert: Zum Einen normalisiert es erhohte TGF-β Spiegel und zum Anderen reduziert L-Carn die Aktivierung des profibrotischen ALK5 Signalweges.